The current project is designed to identify the membrane component(s) responsible for the transport of Na ion, K ion, Ca2 ion and glucose in the human erythrocyte membrane through the use of various protein modifying agents. In addition, the role of cholesterol in alerting these transport processes is also being studied. The compound, 2,3-butanedione, an arginyl specific reagent, has indicated that there is an argenine residue at the binding site of Mg-ATP for the Ca2 ion-dependent ATPase, but not the Mg2 ion-dependent or (Na ion plus K ion)-dependent ATPase systems. The sulfhydryl reagent, N'(iodocetylamino)-1-naphthamine-5-sulfonic acid (1,5 IAEDANS) and the amino specific reagent, Fluorescamine have no effect. The (Na ion plus K ion)-dependent and Ca2 ion-dependent ATPase systems show a marked dependence on the cholesterol content of the human erythrocyte membrane. The inhibition of the (Na ion plus K ion)-dependent ATPase induced by cholesterol modification as completely reversible. However, the inhibition of the Ca2 ion-dependent ATPase system is not reversible. The Mg2 ion- dependent ATPase shows no dependence on the cholesterol content of the erythrocyte membrane. Several estrogen type steroids, ergosterol and 7-dehydrocholesterol protect these ATPase activities against inhibition by preventing cholesterol removal from the membrane. Androgen type steroids do not protect against inhibition.